Profile of Toll-like receptor expressions and induction of nitric oxide synthesis by Toll-like receptor agonists in chicken monocytes

Toll-like receptors (TLRs) play a major role in the innate immune system for initial recognition of microbial pathogens and pathogen associated components. Nitric oxide (NO) is generated in immune cells in response to microbial stimulation and is involved in pathogenesis and control of infection. We used RT-PCR analysis to examine the TLR expression profile on chicken monocytes and demonstrated these cells express chicken TLR2, 3, 4, 6, and 7. TLR5 was not detected by the TR-PCR. We also investigated the differential induction of NO synthesis in chicken monocytes by TLR agonists, including flagellin (FGN, from Salmonella typhimurium), synthetic lipoprotein Pam3CSK4 (PAM), lipopolysaccharide (LPS, from Salmonella enteritidis), lipoteichoic acid (LTA, from Staphylococcus aureus), the synthetic double stranded RNA analog (poly I:C), the guanosine analog, loxoribine (LOX), and synthetic CpG oligodeoxydinucleotide (CpG-ODN). Our results indicate that there was a vast difference among these agonists for their ability to induce NO production. CpG-ODN and LPS were the most potent stimuli and induced significant quantities of NO in cultured monocytes, whereas LTA stimulated significant NO production only at high concentrations. Other agonists such as FGN and poly I:C stimulated very little NO, while PAM, LOX, and nCpG-ODN (control ODN) did not induce NO production. RT-PCR analysis demonstrated that LPS, LTA, and CpG-ODN induced inducible nitric oxide synthase (iNOS) expression in monocytes; whereas the other agonists did not. The presence of TLRs on chicken monocytes and the differential induction of NO production in chicken monocytes by various TLR agonists suggest the differentiation of signaling pathways downstream of individual TLRs.